PROJECT SUMMARY: FUNCTIONAL PROTEOMICS REVERSE PHASE PROTEIN ARRAY (RPPA) CORE The Functional Proteomics Reverse Phase Protein Array (RPPA) Core provides the MD Anderson Cancer Center members and the cancer research community with a powerful, high-throughput, quantitative, cost- effective functional proteomic platform. RPPA measures levels of protein expression and post-translational modification needed for systems biology analysis across large numbers of tumor samples or cell lines. The RPPA Core commenced operations in 2008 with Dr. Yiling Lu as director and recently added Dr. Rehan Akbani as co-director. Current services include protein extraction, robotic arraying of cell/tumor lysates, probing with validated antibodies, and reporting protein expression and modifications to clients. Data are analyzed using customized software (ArrayPro and Supercurve Fitting) to provide signal intensity, curve construction, and relative protein concentration. Importantly, the facility continuously expands the antibody repertoire and has extensively revised its pipeline with emphasis on improved delivery times, quality and accuracy of data. To ensure the utility of the RPPA resource, Drs. Lu and Akbani have multiple active collaborations at MD Anderson, nationally and internationally for their scientific activities and career development. Dr. Lu is the recipient of an NCI Research Specialist (R50) grant, and Dr. Akbani has received 2 U24 grants and an NCI RPPA data generation contract. Since initiating operation, the facility has processed 155,855 samples from 605 investigators (183 cancer center members and 422 external investigators) with between 150 and 302 antibodies. In grant Yr42, 72% of all samples were from peer review-funded users, with the CCSG Breast Cancer and Gynecological Cancers programs both responsible for 25% of the samples. Many of the external users are from other NCI designated cancer centers, including 77 U.S. academic institutions. Over the past 6 years, the RPPA Core has supported 285 publications, with 230 (81%) in journals with IF >5 and 101 (35%) in those with IF >10, including high-profile journals such as N Engl J Med, Nature, and Cell. Furthermore, it provided functional proteomic analysis for 7991 samples from The Cancer Genome Atlas, representing 32 disease types, and the RPPA Core is currently 1 of 2 approved NCI Genomic Characterization Centers. The estimated total operating budget for the current Yr43 is $1,194,149, with 15% ($183,777) from the CCSG. For the past 6-year grant period, MD Anderson provided $1,221,041 of capital equipment support. In the next grant period, $187,727/year (14%) is requested from the CCSG. The RPPA specific aims are: Aim 1) To provide functional proteomic assessment of patient, animal model, and cell line samples to MD Anderson center members using a robust, cost-effective, unbiased antibody-based RPPA approach to analyze the functional status of major signaling pathway networks; Aim 2) To expand the repertoire of validated antibodies for use with RPPA; and Aim 3) To improve quality control of the RPPA data; and Aim 4) To improve the RPPA technology by expanding it to include spatial orientation and quantitative approaches for multiplex analysis.